The present invention is directed to physical structures and methods for separating hematocrit out of small volume whole blood samples leaving merely the plasma or plasma containing a substantially reduced partial volume of hematocrit for testing. The present invention is particularly directed to such structures having no moving parts and employing a capillary region which, subsequent to separation of the plasma, facilitates contacting the reduced hematocrit content plasma by electrodes to permit an accurate detection of an analyte.
Many diagnostic tests are carried out in the clinical field utilizing a blood sample. It is desirable, when possible, to use a very small volumes of blood, often no more than a drop or two. Capillary structures are often employed when handling such small volumes of blood or other fluids. The presence of the hematocrit in the blood sample often interferes with accurate testing and so the removal of, or reduction in concentration of, the hematocrit in the sample, leaving a reduced hematocrit content plasma for testing, is often desirable or even necessary. The removal of the hematocrit is often done using a filter. An example of such a filter device employing capillary structures is described in Hillman, et al., U.S. Pat. Nos. 4,753,776 and 5,135,719. Other devices employing capillary structures to handle whole blood samples are disclosed in McDonald, et al., U.S. Pat. No. 5,039,617; Hillman, et al., U.S. Pat. No. 4,963,498; and Columbus, U.S. Pat. No. 4,271,119.
While such filter devices generally perform satisfactorily, many filter materials tend to absorb a significant portion of the plasma from the blood sample thus leaving only a small volume of the reduced plasma for analytical testing. As the total volume of the sample is diminished, the proportion of the plasma fraction that is absorbed by the filter tends to increase leaving even smaller volumes for testing. It is therefore desirable to construct alternative means for removing hematocrit from whole blood that would be usable on very small sample volumes. It is also desirable to construct such means as will permit testing to be initiated within as short a time span as possible so that the chemistry of the plasma will not have time to change. It is additionally desirable that a number of tests be performed substantially simultaneously on essentially the same sample of blood so that a number of parameters can be measured at the same time.
A capillary hematocrit separation structure according to the present invention is included within a body having an inlet port for receiving a whole blood sample. The body includes a capillary pathway having an inlet end coupled to the inlet port. The capillary pathway is dimensioned sufficiently small to assure transport of the blood sample including the plasma from the inlet end into the capillary pathway by capillary pressure. The capillary pathway includes a plurality of partitions, each partition enclosing a hollow area. Each partition includes at least one opening generally defined by confronting surfaces of the partition. The opening leads from the capillary pathway outside the partition to the hollow area within the partition. The opening dimension defined by the distance between the confronting surfaces of the partition is at least 0.1 xcexcm and generally between about 1.5 and 15 xcexcm so that plasma is encouraged to flow into the hollow area by capillary attraction yet hematocrit is inhibited from passing into the hollow area by virtue of the close proximity of the confronting surfaces of the partition.
In a preferred embodiment, the capillary pathway is defined in part by a first wall and a second wall confronting the first wall. The confronting first and second walls are separated from each other by a distance sufficiently small to assure capillary transport of the blood sample including the plasma between the first and second walls. In this preferred embodiment, the partitions project from the first wall toward the second wall. A plurality of apertures extend through the first wall, each aperture being coincident with a hollow area enclosed by one of the partitions. A backing member is coupled to the first wall so that the backing member and apertures define a plurality of chambers. Each chamber is coupled to one of the hollow areas and includes at least one step surrounding each aperture defining a perimeter of each chamber. The apparatus further comprising a seal surrounding each of the chambers sealing the first wall to the backing member.
A capillary hematocrit separation structure according to the present invention preferably comprises a plurality of pairs of electrodes fixed to the backing member. Each of the electrodes has a first end positioned within one of the chambers and a second end situated adjacent an exposed edge of the backing member. The electrodes are generally arranged in pairs of electrodes so that each chamber includes a first end of two of the electrodes. Preferably, the second ends of all of the electrodes are located on a common exposed edge of the backing member spaced from each other by a regular spacing to permit coupling to an adjacent circuit by a more or less conventional electrical connector.
In the present invention, the average diameter of each hollow area is generally between about 40 and 300 xcexcm, and is typically about 100 xcexcm. The volume of each of the chambers connected to each of the hollow areas is generally between about 0.1 and 100 nl, and is typically about 50 nl. The partitions generally include only a single opening so that there is no flow of liquid through the hollow area enclosed by the partition. The partitions are preferably situated far enough from each other so that their mere proximity to each other does not create a filter effect. The partitions are generally separated from each other by a distance sufficient to ensure that the portion of the sample excluded from any enclosure will have an enhanced partial volume of hematocrit yet still behave as a liquid as it flows into the capillary pathway. Preferably, adjacent partitions are separated from each other, on a nearest neighbor basis, by at least about 10xe2x88x925 meters.
A capillary hematocrit separation structure according to the present invention can be molded as three pieces of a thermoplastic resin such as nylon, styrene-acrylic copolymer, polystyrene, or polycarbonate using known micro-injection molding processes. The mold for making the partitions in the capillary pathway can be constructed by deep reactive ion etching processes typically employed in the manufacture of molds for pre-recorded compact disks and digital video disks. The pieces of the structure are then assembled so that the capillary pathway and the partitions are enclosed within the structure, yet can be accessed at an inlet port designed to receive a sample of blood.
The resulting structure can be viewed as an apparatus for separating a portion of the plasma from a whole blood sample having a selected total volume, the sample including a partial volume of blood plasma and a partial volume of hematocrit. Some fundamental features of the apparatus include a body having an inlet port for receiving a whole blood sample, a capillary pathway having an inlet or proximal end coupled to the inlet port and a distal end including a vent to facilitate fluid flow through the capillary pathway. The capillary pathway is dimensioned sufficiently small to assure transport of blood plasma from the inlet end toward the distal end by capillary pressure. A plurality of partitions are situated in spaced relation from each other on a first wall of the capillary pathway. Each of the partitions surround a hollow area. Each hollow area is connected to the capillary pathway by an opening in the partition dimensioned to inhibit the entry of hematocrit into the hollow area while facilitating the entry of plasma. An aperture is provided in the first wall coincident with each hollow area. Each aperture connects one hollow area to an adjacent chamber. The chamber is defined generally by a relieved region obverse from the adjacent partition and a backing member coupled to the first wall. A first end of a pair of electrodes is positioned within each chamber. The electrodes include a second end positioned on an exposed edge of the apparatus to connect to adjacent circuitry.
The hollow areas within each partition and the adjacent chambers coupled to the hollow areas by the apertures in the first wall form a means for subdividing a whole blood sample into a plurality of much smaller volume samples of plasma containing a reduced concentration of hematocrit. Each of the smaller volume samples is situated in contact with a pair of electrodes which can provide a signal to adjacent circuitry concerning an independent assay conducted on each smaller volume plasma sample. The capillary size and character of the apparatus facilitates the simultaneous testing of a single blood sample for any number of criteria. Any of the test criteria can be duplicated within a given sample to measure the repeatability of any given test, thereby enhancing the confidence attributable to each type of assay. The blood sample is wholly contained within a substantially closed structure formed of low cost materials that facilitate safe disposal of the blood sample following the test.
Other advantageous features will become apparent upon consideration of the following description of a preferred embodiment which references the attached drawings.